This proposal is a competitive renewal for the grant entitled "Genetic Susceptibility to Bladder Cancer: A Molecular Epidemiologic Approach". The purpose of the initial project was to identify interindividual differences in susceptibility to tobacco-induced carcinogenesis, as risk factors for bladder cancer (BC). We now propose to build upon this epidemiologic and specimen resource of 600 cases and 600 controls and to recruit an additional 400 patients with newly diagnosed BC and 400 healthy controls matched to the cases on the basis of sex, age (+/- 5 years), and ethnicity. We intend to apply promising novel phenotypic biomarkers of constitutive genetic instability, such as telomere length and mutagen-induced DNA damage. Specifically, we plan to determine telomere length in peripheral blood lymphocytes (PBLs) of the newly recruited 400 BC patients and 400 controls using quantitative fluorescent in situ hybridization laser scanning cytometry (Q-FISH/LSC). Our hypothesis is that BC cases exhibit shortened telomeres compared with control subjects. To assess latent genetic instability, we plan to quantify mutagen-induced DNA damage using the comet assay in the PBLs of 400 BC patients and 400 controls. Our hypothesis is that BC patients exhibit increased levels of BPDE-induced and or gamma-radiation-induced genetic damage. To evaluate the role of genetic variations, we will use a pathway approach to estimate the frequencies of single-nucleotide polymorphisms (SNPs) in DNA repair genes implicated in the nucleotide excision repair (NER), base excision repair (BER), and double-stranded break (DSB) pathways for all 1000 cases and 1000 controls. Our hypothesis is that individuals with adverse genotypes of the NER, BER, and DBS pathways are at increased risk for BC and that these genotypes predict higher levels of induced DATA damage. To assess whether surrogate (PBL) tissue reflects molecular events in the target tissue, we plan to determine telomere length in paired samples of urine epithelial cells and blood lymphocytes from 75 patients with superficial BC and 75 healthy controls. Our hypothesis is that there will be a positive correlation in telomere length between target and surrogate tissues. Finally, using a multi-color fluorescent in situ hybridization (FISH) assay that includes a mixture of probes for the centromeric regions in chromosomes 3, 7, and 17 and the 91321 region, we will test the hypothesis that individuals with short telomeres, adverse genotypes and or high mutagen-induced DNA damage in PBLs (surrogate tissue) exhibit higher levels of genetic instability in the target tissue. As a secondary aim, we will correlate the predicted functional significance of these SNPs using a computational algorithm approach with risk estimated from this proposal to develop new tools for future candidate SNPs selection. The ability to rapidly screen individuals for risk, using minimally invasive procedures (blood samples), has immense clinical implication, such as intensive screening and chemopreventive interventions.